Large-scale production of recombinant proteins, such as antibodies, typically relies on expression of the protein from a cultured production cell line. Proteins expressed by cultured cells can be readily recovered and purified using standard bench or industrial-scale purification techniques.
Antibody-producing cell lines can be created by random integration of heavy chain and light chain antibody expression constructs into host cell genomic DNA. This random integration yields a high percentage of stable clones having relatively low levels of antibody expression. This phenomenon may be due to the fact that many integrations insert transgenes into areas of condensed chromatin, which are not conducive to high levels of expression (Furth et al., Nucl. Acids. Res., 19, 6205-6208 (1991)).
Protein expression level is an important parameter affecting the production and purification of such proteins from a bioreactor or other system. In general, higher purified protein yields can be attained when the expression level is relatively high. Conversely, if the expression level is low, yields may be low and protein purification may not be economically feasible.
One approach to circumventing the problem of cells with low protein expression has been to subclone and isolate high expressing cells from a population of low expressing cells. Typically, this requires several time and labor-intensive rounds of limiting serial dilution, screening and selection of high expressing cell lines. Alternatively, entirely new cell lines producing the protein are generated with the intent that the new cell lines will be high expressing lines.
However, neither of these approaches offer any guarantee of success and both have significant limitations. For example, identifying high expressing cell lines by subcloning from a population of low expressing cells is limited by the relative rarity of high expressing cells in the population as well as the extensive amounts of time and labor required for the identification of any high expressing cells. Further, new cell lines producing the antibody or protein of interest may not continue to be high expressing, due to silencing of the transgene expression during expansion and growth of the cell line or other forms of genetic or epigenetic instability. (Insulators would not help)
Thus, a need exists for compositions, methods, and cells that effectively increase peptide chain expression.